The goal of this project is to clone the high affinity choline uptake carrier (HACU), an important molecular reagent for understanding cholinergic mechanism during normal aging process and under several pathological conditions such as Alzheimer's disease and Amyotrophic Lateral Sclerosis (ALS). Cholinergic neurotransmission in the central nervous system is involved in many fundamental brain processes such as learning, arousal and sleep, which undergo age related alterations. The formation of the neurotransmitter acetylcholine is catalyzed by the enzyme choline acetyltransferase (ChAT, E.C. 2.3.1.6), which transfers acetyl group from acetylcoenzyme A to choline, in the presynaptic nerve terminals of cholinergic neurons. Acetylcholine is packaged into the synaptic vesicles by a vesicular acetylcholine transporter (VAChT) and is then ready to be released in a calcium dependent manner. Acetylcholine binds specifically to either the nicotinic or muscarinic receptors to transmit information to the postsynaptic neurons. The action of acetylcholine is terminated through hydrolysis to acetate and choline by the enzyme acetylcholinesterase (ACHE, E.C.3.1.1.7). Most of the choline is then transported back to the presynaptic terminal to be recycled as one of the precursors for the biosynthesis of the neurotransmitter acetylcholine. This step, which is mediated by the action of the high affinity choline uptake system (HACU), is believed to be the rate limiting step in the biosynthesis of acetylcholine. Therefore, molecular analysis of the HACU is crucial for further understanding of how cholinergic mechanism operates in the central nervous system and is regulated under physiological and pathological conditions.